The general aim of this project is the elucidation of the mechanism of action of cardiac glycosides at the cellular and the molecular levels. Attention is focused on the interaction of these drugs with Na ion, K ion-ATPase of the cell membrane. The specific projects proposed are: 1. Using the fluorimetric assay of methylfluorescine phosphatase for the determination of extent of inhibition of enzyme in drug-exposed dog and rabbit hearts, the relation of positive inotropic effects of cardiac glycosides to their inhibitory effects on Na ion, K ion-ATPase will be re-examined. 2. Through studies on the kinetics of cardiac glycoside interaction with the enzyme of intact human red cells, the determination of the mechanism of effect of extracellular K ion on this drug-receptor interaction; and the elucidation of the cause of extreme stability of this drug receptor complex in the intact cell, will be attempted. 3. Effects of cardiac glycosides on enzymes prepared from Purkinje fibers of the hearts of newborn and adult dogs will be studied, to determine if different drug-sensitivities of these enzymes can account for the different responses of these hearts to cardiac glycosides. 4. To learn more about the molecular bases of different sensitivities of various purified enzymes to cardiac glycosides, the properties of three enzymes with known differences in drug sensitivities will be compared. These are the native dog kidney enzyme, the native rat kidney enzyme, and the dog kidney enzyme whose ATP-regulatory site has been blocked through reaction with ethylmercury.